Since gamma-interferon (IFN-gamma) is a potent immunomodulator and patients receiving certain antineoplastic agents are at risk of unusual infections, we have determined the effect of certain antineoplastic agents on IFN-gamma production. Induction of peripheral blood mononuclear cells from normal donors in the presence and absence of various antineoplastic agents was achieved using phytohemagglutinin (8 micrograms/ml). Supernatants were then separated by centrifugation, dialyzed, and assayed for interferon. Cell viability was always greater than 85% with or without the presence of drugs. Hydrocortisone was found to eliminate IFN-gamma production if added within 24 hr after the phytohemagglutinin. The suppression of IFN-gamma production occurred with hydrocortisone concentrations as low as 0.65 microgram/ml, was associated with a diminished proliferative response to the lectin, and occurred with other interferon inducers including staphylococcal enterotoxin A. Adriamycin (0.4 microgram/ml) and vincristine (0.08 microgram/ml) also diminished IFN-gamma production, but only if the peripheral blood mononuclear cells were pretreated with the drugs. Methotrexate, 5-fluorouracil, and 6-mercaptopurine failed to influence the yield of IFN-gamma. These results are significantly different from experiments previously reported using alpha- and beta-interferons and suggest an important mechanism by which these drugs can produce immunosuppression.